1. Field of the Invention
The present invention relates to a histochemical stain, a method for producing the stain, and a method for the post-mortem detection and localization of myelin in brain tissue slices.
2. Description of the Prior Art
On the most fundamental level, the brain can be divided into two distinctly different appearing types of matter, the white matter and the gray matter. In recent years, numerous histochemical techniques have been developed for the localization of neuronal and astrocytic markers within the gray matter. Fewer markers, however, are available to the researcher studying the myelin which comprises the white matter. Existing methods for staining myelin include those based on lipid solubility such as Sudan black or Oil red O, the affinity for non-solvent extractable phospholipids by Luxol Fast Blue, the chelation of complex lipid polymers with potassium dichromate followed by hematoxylin, the suppression of non-myelin argyrophilia with pyridine followed by diamine silver, the immunohistochemical localization of myelin basic protein, and the use of aqueous gold chloride solutions.
The use of gold salts as a myelin stain has a long and controversial history. The use of gold chloride as a sensitive myelin stain is known in the art. The use of gold chloride in hypotonic phosphate buffered saline to study the myelinopathy resulting from exposure to isoniazid or fumonisin indicates that although the gold chloride based method has the potential to detect myelin pathologies, it, like the previous gold chloride based methods, also suffered from a certain degree of capriciousness.
The principal object of the present invention is to provide a stable, soluble, gold based complex stain which retains the advantages associated with the traditional gold chloride stains and methods while eliminating the drawbacks associated therewith.
Another object of the invention is to provide a stain of the foregoing character for simply and sensitively labeling normal and pathological myelin in brain tissue sections.
A further object of the invention is to provide a stain of the foregoing character which is consistent in results and affords relatively short staining times.
In accordance with the foregoing objects, the novel histochemical marker or stain embodying the present invention was synthesized and was applied as a stain or label to demonstrate both normal and pathological myelin. This stain is identified as a aurophosphate complex, more specifically a potassium aurophosphate, [K6Au(PO4)3]n, produced as the reaction product of dibasic potassium phosphate and a gold chloride.
To examine pathological myelin changes, a number of agents known to cause brain damage were used including isoniazid, 3-nitropropionic acid, kainic acid, domoic acid, and intracranial injection of amidino stilbene.